PROJECT SUMMARY BMD (bone mineral density) is the most commonly studied major risk factor for osteoporosis (OP). So far, GWAS studies have identified ~100 BMD marker loci with common variants, in samples with females only or with both males and females. The identified loci, in total, explain <10% BMD heritability and their sex-specific genetic effects are largely unknown. In addition, specific functional DNA sequence variants are also generally unknown. The Goal of this project is to most comprehensively identify potential functional DNA sequence variants, in particular those rare and structural variants, for male BMD, and to assess their sex/ethnicity generality/specificity and their ultimate importance to the most devastating clinical outcome of OP - osteoporotic hip fracture (HF). During the past ~two decades, we have 1) recruited one of the largest samples in the field; 2) established broad collaboration; 3) assembled an experienced team with extensive multi-disciplinary research productivity. In this project, we propose to capitalize on our unique extensive resources to fulfill the following Specific Aims: Aim 1. Identify (Discovery) potential functional variants for male OP risk by whole-genome sequencing We will perform a large-scale whole-genome sequencing (WGS) to search for functional variants associated with BMD in 3,200 unrelated US Caucasian males with extremely discordant hip BMDs (1,600 with high and 1,600 with low hip BMD). To further increase the power, we will perform WGS imputation (for common & rare variants) in >6,500 independent male subjects (used in our earlier GWAS) for combined analyses with the WGS data. Aim 2. Validate promising functional variants discovered in Aim 1 and identify those sex-specific ones We will validate the 200 top variants identified above in an independent sample of 12,000 US Caucasians (both males and females). Sex-specific or sex-general effects of these variants will be assessed. Aim 3. Assess ethnicity/population-generality/specificity of the identified variants Ethnicity/population-generality/specificity of the top ~50 most significant variants validated in Aim 2 will be tested in ~85,000 subjects, including Caucasians, African Americans, Hispanic Americans, and East Asians. Aim 4. Test the identified BMD variants for their relevance to HF We will test at least 20 top BMD variants validated in Aim 2 for their relevance to HF, in the world largest collection of an independent sample of 10,493 HF cases and 10,815 matched controls. Data from this project will be examined integratively with other functional omics data from Proj 2&3 (transcriptome and epigenome) in this U19 program project and from our proteomics project (R01AR057049, PI Dr. Deng). The purposes are to 1) gain insights into the potential functional mechanisms of the variants/genes identified in this project, 2) identify additional significant functional variants/genes by synthesizing various omics data. Regular eQTL (expression QTL), mQTL (methylation QTL), pQTL (protein expression QTL) analyses and innovative integrative multi-omics analyses will be performed in Proj 1-3 and particularly in Biostatistics and Bioinformatics Core.